Necrosis is a major contributor to human disease, however little effort has been made to develop specific therapies targeting pathologic necrosis due it its perceived uncontrolled nature. This notion has recently been challenged by the discovery that intrinsic death pathways down stream of tumor necrosis factor receptor family members can initiate necrotic death, suggesting that necrosis can be specifically targeted for inhibition. We have recently developed necrostatins, potent and selective small molecule inhibitors of necroptosis, a form of programmed necrosis initiated by TNF alpha and Fas receptor. Necrostatin-1 was used to directly establish necroptosis as a major component of neuronal cell death after ischemic brain injury in vivo. Traumatic brain injury is a leading cause of death and life-long disability in young adults and children, however mechanisms of traumatic brain cell death and associated neurological dysfunction are not well characterized. We have shown that TNF and Fas play a key role in histopathological and functional outcome after controlled cortical impact (CCI) in mice. Since TNF and Fas are well established inducers of necroptosis, we investigated whether necroptosis contributes to outcome after TBI. We found that necrostatin-1 reduced tissue damage and markedly improved functional outcome following CCI in mice. These results suggest that the target of necrostatin-1, receptor interacting protein kinase 1 (RIPK1), may represent an exciting new target for TBI therapy. To further investigate the role of RIPK1 in TBI, we propose three Specific Aims: 1) Establish the role of RIPK1 in neuronal cell death in a variety of in vitro neuronal death paradigms related to TBI using knockdown (RIPK1 shRNA) and dominant negative (RIPK1 K45M) strategies; 2) Demonstrate a key contribution of RIPK1 to brain trauma in vivo, using AAV-8 viral vector transduction of neurons in vivo with shRNA RIPK1 or RIPK1 K45M constructs; and 3) Determine RIPK1-related signaling pathways operative during necroptosis in cultured neurons and in injured brain after TBI in vivo. PUBLIC HEALTH RELEVANCE: Work in the proposed project could render a new treatment for patients with traumatic brain injury. The proposed drug therapy might reduce brain cell death as well as improve learning and memory in survivors, and thereby improve their quality of life.